Identification, Phylogeny, and Expression Profiling of Pineapple Heat Shock Proteins (HSP70) Under Various Abiotic Stresses

Abstract

Pineapple (Ananas comosus (L.) Merr.) is an economically significant and delicious tropical fruit. Pineapple commercial production faces severe decline due to abiotic stresses, which affect the development and quality of pineapple fruit. Heat shock protein 70 (HSP70) plays an essential role in abiotic stress tolerance. However, the pineapple HSP70 family identification and expression analysis in response to abiotic stresses has not been studied. To explore the functional role of AcHSP70, different abiotic stress treatments were applied to pineapple cultivar "Bali" seedlings. A total of 21 AcHSP70 members were identified in the pineapple genome. The identified genes were classified into four subfamilies (I-IV) using phylogenetic analysis. The AcHSP70 family is expressed under different stress conditions. Quantitative real time polymerase chain reaction (qRT-PCR) revealed the expression pattern of the AcHSP70 family under cold, drought, salt, and heat stress. The expression level of genes such as AcHSP70-2 increased under heat, cold, and drought stress, while the expression level of genes such as AcHSP70-3 decreased under salt stress. Furthermore, the expression profile of AcHSP70s in different tissues and development stages was analyzed using transcriptome analysis. The HSP70 genes exhibited unique expression patterns in pineapple tissue at different developmental stages. The study therefore provides a list of HSP70 genes with substantial roles in abiotic stress response and valuable information for understanding AcHSP70 functional characteristics during abiotic stress tolerance in pineapple.

Keywords: AcHSP70 family; Ananas comosus; abiotic stress; gene expression; phylogeny.

Figure 1

Chromosome distribution of AcHSP70 genes in pineapple. The AcHSP70s were located on Chr 2, 3, 4, 7, 8, 13, 14, 16, 17, 19, 20, 21, 22, and 25. Chr: chromosome. The ruler located on the left side represents the chromosome length and is shown in megabase (Mb).

Figure 2

The phylogenetic analysis of AcHSP70 proteins with Arabidopsis, cucumber (Cucumis sativus L.), rice (Oryza sativa L.), and maize (Zea mays L.). The phylogenetic tree was made by using MEGA 11.0 software with the neighbor-joining (NJ) method, and the bootstrap replications were set to 1000 times. Different colors represent four groups (I–IV), and stars represent AcHSP70s.

Figure 3

Gene structures and conserved motifs of AcHSP70s. (A) Different colors represent the four groups of AcHSP70 genes (I–IV). (B) The motifs of AcHSP70 proteins are shown as colored boxes. (C) Gene structures of AcHSP70 genes. The yellow blocks represent the coding sequence (CDS), the green blocks represent the untranslated region (UTR), and the black lines represent introns.

Figure 4

Three-dimensional structural analysis of AcHSP70s.

Figure 5

The cis-acting elements in promoters of AcHSP70 genes. The amounts of cis-elements in AcHSP70s promoter regions were displayed in different colors and numbers in the grid.

Figure 6

Intraspecies synteny analysis of AcHSP70 genes. The black curve represents duplication events between AcHSP70 genes. Chr 1–25: Chromosome 1–25.

Figure 7

Collinearity of HSP70 genes in pineapple. The gray line represents the collinearity of all the genes in the pineapple, and the red line represents the collinearity of the AcHSP70 genes.

Figure 8

Expression profiles of AcHSP70 family members in pineapple leaves with and without spines. Transcriptomic data (Le_1: Leaf apices; Le_2: Leaf base; Ro: Root; Fl: Flower; Fr: fruit) were analyzed using Log2(FPKM) values. The color scale on the right represents the relative expression level, from high (orange) to low (blue).

Figure 9

Expression levels of AcHSP70 genes under 0 h control (CK), 4 h, 12 h, 24 h, and 72 h of heat stress treatment. Data are expressed as means ± SD (n = 3). Different letters indicate significant differences between groups (p < 0.05).

Figure 10

Expression levels of AcHSP70 genes under 0 h control (CK), 4 h, 12 h, 24 h, and 72 h of cold stress treatment. Data are expressed as means ± SD (n = 3). Different letters indicate significant differences between groups (p < 0.05).

Figure 11

Expression levels of AcHSP70 genes under 0 h control (CK), 4 h, 12 h, 24 h, and 72 h of drought treatment. Data are expressed as means ± SD (n = 3). Different letters indicate significant differences between groups (p < 0.05).

Figure 12

Expression levels of AcHSP70 genes under 0 h control (CK), 4 h, 12 h, 24 h, and 72 h of salt stress treatment. Data are expressed as means ± SD (n = 3). Different letters indicate significant differences between groups (p < 0.05).